Redefining ITT cortisol thresholds on Abbott platforms to prevent misdiagnosis of adrenal insufficiency.

BACKGROUND: Adrenal insufficiency (AI) is a life-threatening condition which requires long term glucocorticoid replacement. The insulin tolerance test (ITT) is the current gold standard test for diagnosis of secondary AI, but the widely accepted cut-off value of a peak cortisol of less than 500 nmol/L assumes that anyone who does not reach this value has AI and thus requires full replacement. The cut-off used to diagnose AI is also founded on outdated assays. Use of this cut-off in an era of more specific immunoassays therefore risks misdiagnosis, subsequent unnecessary glucocorticoid exposure and associated adverse effects with increased mortality risk. DESIGN, PATIENTS AND MEASUREMENTS: This retrospective analysis assessed 300 ITT cortisol responses using the Abbott Architect and Alinity analyser platforms in patients with suspected AI over a period of 12 years (August 2010 to January 2022), at a tertiary centre. RESULTS: Patients were classified as having AI or not, based on a comprehensive clinical review of electronic patient records from the point of test to the present day by a panel of pituitary and adrenal specialists. Using the current institutional cut-off value of 500 nmol/L, receiver operating characteristic analysis identified a 100.0% sensitivity and 43.6% specificity (area under the curve 0.979). Using a lower cortisol threshold value of 416 nmol/L on the Abbott analyser platform maintained a sensitivity of 100.0% and improved the specificity to 86.7%. CONCLUSION: This data supports lowering the Abbott analyser ITT peak cortisol threshold to 416 nmol/L. Use of this improved cut-off avoids unnecessary glucocorticoid replacement therapy in 104 (34.7%) of individuals in this study. All patients remained well with at least 1 year longitudinal follow up of glucocorticoid replacement.

Sex steroid priming for growth hormone stimulation testing in children and adolescents with short stature: A systematic review.

OBJECTIVE: Growth hormone stimulation testing (GHST) is used to diagnose growth hormone deficiency (GHD) in children. As sex steroids impact on anterior pituitary function, there is concern around the efficacy of GHST in peripubertal children, where endogenous sex steroid levels are low. Sex steroid priming before GHST is thought to improve test efficacy in these children, however evidence to support its use in clinical practice is limited. In this systematic review, we addressed the following research questions: Does priming increase GH stimulation test efficacy in peripubertal children? Does priming identify those who would benefit most from treatment in terms of final height? Is there evidence for an optimal sex-steroid priming regimen? DESIGN, PATIENTS, MEASUREMENTS: The study was registered with PROSPERO and conducted according to PRISMA guidelines. We searched Medline, Cochrane-Library, Scopus, EMBASE and Web-of-Science and included all studies that included GHST in both primed and unprimed children. A GH cut-off of 7 µg/L was used as a threshold for GHD. Study quality was assessed using the Risk-Of-Bias in Non- Randomized Studies (ROBINS-I) tool or the revised Cochrane risk-of-bias tool for Randomised trials. RESULTS: Fifteen studies met our inclusion criteria, of which 4/15 (27%) were randomised control trials. The majority (9/15) of the studies indicated that priming increases growth hormone response upon GHST in peripubertal children, increasing test specificity. Two studies investigated final height after treatment based on the results of primed versus unprimed GHST. These results indicate that growth hormone treatment based on results of a primed GHST improve outcomes compared with treatment based on an unprimed test. CONCLUSION: Sex-steroid priming increases the growth hormone response during GHST, resulting in fewer patients meeting the threshold required for a diagnosis of GHD. Unnecessary GH treatment may be avoided in some patients without a detrimental effect on final height. Numerous sex-steroid priming regimens have been used in clinical practice and the majority appear to be effective, but an optimal regimen has not been determined.

The Characterization of Sex Differences in Hypoglycemia-Induced Activation of HPA Axis on the Transcriptomic Level.

Activation of the hypothalamic-pituitary-adrenal (HPA) axis using an insulin tolerance test (ITT) is a medical diagnostic procedure that is frequently used in humans to assess the HPA and growth-hormone (GH) axes. Whether sex differences exist in the response to ITT stress is unknown. Thus, investigations into the analysis of transcripts during activation of the HPA axis in response to hypoglycemia have revealed the underlying influences of sex in signaling pathways that stimulate the HPA axis. We assessed four time points of ITT application in Balb/c mice. After insulin injection, expression levels of 192 microRNAs and 41 mRNAs associated with the HPA, GH and hypothalamic-pituitary-gonadal (HPG) axes were determined by real-time RT-PCR in the hypothalamus, pituitary and adrenal tissues, as well as blood samples (Raw data accession: https://drive.google.com/drive/folders/10qI00NAtjxOepcNKxSJnQbJeBFa6zgHK?usp=sharing ). Although the ITT is commonly used as a gold standard for evaluating the HPA axis, we found completely different responses between males and females with respect to activation of the HPA axis. While activation of several transcripts in the hypothalamus and pituitary was observed after performing the ITT in males within 10 min, females responded via the pituitary and adrenal immediately and durably over 40 min. Additionally, we found that microRNA alterations precede mRNA responses in the HPA axis. Furthermore, robust changes in the levels of several transcripts including Avpr1b and Avpr2 observed at all time points strongly suggest that transcriptional control of these genes occurs mostly via differential signaling in pituitary and blood between males and females. Male and female HPA axis responses to ITT involve a number of sophisticated regulatory signaling pathways of miRNAs and mRNAs. Our results highlight the first robust markers in several layers of HPA, HPG and GH axis involved in ITT/hypoglycemia stress-induced dynamics.

Clinical characteristics of patients with unexplainable hypothalamic disorder diagnosed by the corticotropin-releasing hormone challenge test: a retrospective study.

BACKGROUND: The corticotropin-releasing hormone (CRH) challenge test can distinguish the disorders of the hypothalamus from those of the pituitary. However, the pathophysiology of hypothalamic disorder (HD) has not been fully understood. This study aimed to elucidate the clinical characteristics of patients with unexplainable HD, diagnosed by the CRH challenge test. METHODS: We retrospectively reviewed patients who underwent the CRH challenge test. Patients were categorized into four groups as follows: patients with peak serum cortisol ≥18 µg/dL were assigned to the normal response (NR) group (n = 18), among patients with peak serum cortisol < 18 µg/dL and peak adrenocorticotropic hormone (ACTH) increase ≥two-fold, patients without obvious background pathology were assigned to the unexplainable-HD group (n = 18), whereas patients with obvious background pathology were assigned to the explainable-HD group (n = 38), and patients with peak serum cortisol < 18 µg/dL and peak ACTH increase

Wheel running leads to sex-specific effects on Western diet-associated glucose homeostasis and brain insulin signaling without altering food-related impulsive choice.

OBJECTIVES: There is a clear association between obesity and impulsivity. While exercise can suppress weight gain and decrease impulsive choice (IC), the relationship between impulsivity, the consumption of palatable, energy dense diets, and exercise is unclear. We examined IC before and after Western diet (WD) exposure in rats of both sexes and whether exercise would rescue any diet-mediated increases in IC. Our hypotheses were twofold: first, increased impulsivity would be associated with higher WD preference in a positive feedback loop and second, increased WD consumption would impair both peripheral and central insulin signaling, both of which exercise would attenuate. METHODS: Following baseline assessment of IC through a delay discounting task, rats were divided into naïve, sedentary (Sed), or wheel running (WR) groups for a 5-week WR and two-diet choice period after which rats underwent an oral glucose (OGTT) and insulin tolerance test (ITT) in addition to a re-test of IC. Insulin induced Akt-GSK3ß signaling in the brain was examined using western blot. RESULTS: All Sed rats preferred the WD diet, and all WR rats initially avoided the WD but subsequently reversed their avoidance to preference with females reversing earlier than males. Exercise suppressed weight gain and adiposity to a greater extent in males than females. Only WR males showed improved glucose clearance during OGTT, but both male and female WR rats had a faster recovery of hypoglycemia during ITT. Furthermore, WR rescued WD-induced deficits in hypothalamic Akt-GSK3ß signaling in males but not females. In the prefrontal cortex, however, WD and WR both reduced Akt-GSK3ß signaling in males but not females. There were no sex differences in IC at baseline, and all rats made more impulsive choices during the re-test independent of diet, sex, or exercise. DISCUSSION: The results suggest that while exercise may have a greater efficacy at attenuating diet-mediated metabolic dysregulation in males, it has some beneficial effects for females and highlights the need to develop sex-specific interventions for restoring energy balance.

Is the fear from insulin tolerance test in the evaluation of short stature justified?

Among growth hormone (GH) provocative tests, insulin tolerance test (ITT) is considered a valuable test in children with short stature to diagnose GH deficiency. However, many pediatric endocrinologists are reluctant to perform it. We conducted a nationwide survey among all the pediatric endocrine clinics' heads in Israel regarding their position on this issue. We found that the number of endocrine units performing ITT is almost nil. Sense of fear from severe hypoglycemia was a dominating cause for not performing ITT. We review the pros and cons of performing ITT versus other tests, especially glucagon test. Glucagon tolerance test is not considered the test of choice by some endocrinologists but recommended by others including in Israel. We also note the influence of a widely cited report published in 1992 (many times inaccurately) on endocrinologists' views. CONCLUSION: A nationwide survey in Israel revealed a high rate of reluctance to perform ITT. The rationale behind this attitude was a sense of fear of performing the test by many endocrinologists. We discuss the preferences for choosing GH stimulation tests and the pros and cons of alternatives to ITT. The fear of not performing ITT was not always justified. WHAT IS KNOWN: • ITT test is considered a gold standard in the evaluation of short stature to diagnose GH deficiency by many endocrinologists. WHAT IS NEW: • High reluctance rate found in Israel to perform ITT, prompted us to evaluate the attitudes of pediatric endocrinologists around the world showing conflicting ideas. • The role of a single paper sometimes misquoted also contributed to these conflicting results.

Strain-based and sex-biased differences in adrenal and pancreatic gene expression between KK/HlJ and C57BL/6 J mice.

BACKGROUND: The ever-increasing prevalence of diabetes and associated comorbidities serves to highlight the necessity of biologically relevant small-animal models to investigate its etiology, pathology and treatment. Although the C57BL/6 J model is amongst the most widely used mouse model due to its susceptibility to diet-induced obesity (DIO), there are a number of limitations namely [1] that unambiguous fasting hyperglycemia can only be achieved via dietary manipulation and/or chemical ablation of the pancreatic beta cells. [2] Heterogeneity in the obesogenic effects of hypercaloric feeding has been noted, together with sex-dependent differences, with males being more responsive. The KK mouse strain has been used to study aspects of the metabolic syndrome and prediabetes. We recently conducted a study which characterized the differences in male and female glucocentric parameters between the KK/HlJ and C57BL/6 J strains as well as diabetes-related behavioral differences (Inglis et al. 2019). In the present study, we further characterize these models by examining strain- and sex-dependent differences in pancreatic and adrenal gene expression using Affymetrix microarray together with endocrine-associated serum analysis. RESULTS: In addition to strain-associated differences in insulin tolerance, we found significant elevations in KK/HlJ mouse serum leptin, insulin and aldosterone. Additionally, glucagon and corticosterone were elevated in female mice of both strains. Using 2-factor ANOVA and a significance level set at 0.05, we identified 10,269 pancreatic and 10,338 adrenal genes with an intensity cut-off of ≥2.0 for all 4 experimental groups. In the pancreas, gene expression upregulated in the KK/HlJ strain related to increased insulin secretory granule biofunction and pancreatic hyperplasia, whereas ontology of upregulated adrenal differentially expressed genes (DEGs) related to cell signaling and neurotransmission. We established a network of functionally related DEGs commonly upregulated in both endocrine tissues of KK/HlJ mice which included the genes coding for endocrine secretory vesicle biogenesis and regulation: PCSK2, PCSK1N, SCG5, PTPRN, CHGB and APLP1. We also identified genes with sex-biased expression common to both strains and tissues including the paternally expressed imprint gene neuronatin. CONCLUSION: Our novel results have further characterized the commonalities and diversities of pancreatic and adrenal gene expression between the KK/HlJ and C57BL/6 J strains as well as differences in serum markers of endocrine physiology.

Investigation of the Hypothalamo-pituitary-adrenal (HPA) axis: a contemporary synthesis.

The hypothalamo-pituitary-adrenal (HPA) axis is one of the main components of the stress system. Maintenance of normal physiological events, which include stress responses to internal or external stimuli in the body, depends on appropriate HPA axis function. In the case of severe cortisol deficiency, especially when there is a triggering factor, the patient may develop a life-threatening adrenal crisis which may result in death unless early diagnosis and adequate treatment are carried out. The maintenance of normal physiology and survival depend upon a sufficient level of cortisol in the circulation. Life-long glucocorticoid replacement therapy, in most cases meeting but not exceeding the need of the patient, is essential for normal life expectancy and maintenance of the quality of life. To enable this, the initial step should be the correct diagnosis of adrenal insufficiency (AI) which requires careful evaluation of the HPA axis, a highly dynamic endocrine system. The diagnosis of AI in patients with frank manifestations is not challenging. These patients do not need dynamic tests, and basal cortisol is usually enough to give a correct diagnosis. However, most cases of secondary adrenal insufficiency (SAI) take place in a gray zone when clinical manifestations are mild. In this situation, more complicated methods that can simulate the response of the HPA axis to a major stress are required. Numerous studies in the assessment of HPA axis have been published in the world literature. In this review, the tests used in the diagnosis of secondary AI or in the investigation of suspected HPA axis insufficiency are discussed in detail, and in the light of this, various recommendations are made.

The Cut-off Limits of Growth Hormone Response to the Insulin Tolerance Test Related to Body Mass Index for the Diagnosis of Adult Growth Hormone Deficiency.

INTRODUCTION: The diagnosis of growth hormone deficiency (GHD) in adults is based on a reduced GH response to provocative tests, such as the insulin tolerance test (ITT) and the GH-releasing hormone (GHRH) + arginine (ARG) test. However, the cut-off limits of peak GH response in lean subjects are not reliable in obese patients; this is noteworthy since adult GHD is often associated with obesity. To date, there are no ITT cut-offs related to body mass index (BMI). OBJECTIVE: We aimed to evaluate the diagnostic cut-offs of GH response to the ITT in the function of BMI. METHODS: The GH response to the ITT was studied in 106 patients with a history of hypothalamic-pituitary disease, a mean age of 48.2 ± 12.4 years, and a mean BMI of 26.8 ± 6.1 kg/m2). Patients were divided into lean, overweight, and obese groups according to their BMI. The lack of GH response to GHRH + ARG test was considered the gold standard for the diagnosis of GHD. The best GH cut-off in the ITT, defined as the one with the best sensitivity (SE) and specificity (SP), was identified using receiver-operating characteristics curve (ROC) analysis. RESULTS: The best GH cut-off in the ITT was 3.5 µg/L in lean subjects (SE 82.1%; SP 85.7%), 1.3 µg/L in overweight subjects (SE 74.1%; SP 85.7%), and 2.2 µg/L in obese subjects (SE 90.0%; SP 50.0%). The diagnostic accuracy was 97.2, 76.5, and 76.7%, respectively. CONCLUSIONS: Our data show that the ITT represents a reliable diagnostic tool for the diagnosis of adult GHD in lean subjects if an appropriate cut-off limit is assumed. Overweight and obesity strongly reduce the GH response to the ITT, GH BMI-related cut-off limits, and the diagnostic reliability of the test.

Morning Serum Cortisol Level Predicts Central Adrenal Insufficiency Diagnosed by Insulin Tolerance Test.

INTRODUCTION: According to guidelines, a morning serum cortisol level <83 nmol/L is diagnostic for central adrenal insufficiency (CAI), a value >414 nmol/L excludes CAI, while values between 83 and 414 nmol/L require stimulation tests. However, there are no currently reliable data on morning serum cortisol for prediction of cortisol response to insulin tolerance test (ITT). OBJECTIVE: Using the receiver-operating characteristic curve analysis, the purpose of this study was to detect the morning serum cortisol cutoff with a specificity (SP) or a sensitivity (SE) above 95% that identify those patients who should not be tested with ITT. METHODS: We included 141 adult patients (83 males) aged 42.7 ± 12.3 (mean ± standard deviation) years old. Based on the serum cortisol response to ITT, patients have been divided into 2 groups: subjects with CAI (peak serum cortisol <500 nmol/L; 65 patients) and subjects with preserved adrenocortical function (peak cortisol >500 nmol/L; 76 patients). RESULTS: The best morning cortisol cutoff, in terms of SE (87.7%) and SP (46.1%), was ≤323.3 nmol/L. The cutoff of morning serum cortisol concentration that best predicted a deficient response to ITT was ≤126.4 nmol/L (SE 13.8%, SP 98.7%). The cutoff of morning serum cortisol concentration that best predicted a normal response to ITT was >444.7 nmol/L (SE 96.9%, SP 14.5%). CONCLUSIONS: This is the first study that identifies a morning serum cortisol cutoff that best predict the response to ITT in order to simplify the diagnostic process in patients with suspected CAI. A new diagnostic flow-chart for CAI is proposed.

Revisiting peak serum cortisol response to insulin-induced hypoglycemia in children.

PURPOSE: To evaluate factors that could potentially affect the hypothalamic-pituitary adrenal (HPA) axis response to insulin-induced hypoglycemia in children without history or symptoms of adrenal insufficiency and to propose a cut-off value to define a normal response in this population. METHODS: Exploratory single-center study involving 78 children that prospectively underwent insulin tolerance test (ITT) for suspected growth hormone (GH) deficiency. METHODS: Glucose, cortisol, GH, adrenocorticotrophic hormone (ACTH), epinephrine and norepinephrine levels were measured at baseline and after insulin-induced hypoglycemia. Serum cortisol was measured using Access automated immunoassay. RESULTS: Mean (range) basal morning serum cortisol of 8 (2.2-19.5) µg/dL/222 (61-542) nmol/L increased after hypoglycemia to 20.5 (14.6-29.5) µg/dL/570 nmol/L (405-819) nmol/L. Peak serum cortisol levels of 14.6 µg/dL (405 nmol/L) and 15.4 µg/dL (428 nmol/L) corresponded to the 2.5th and 5th percentiles, respectively. Peak serum cortisol correlated with peak plasma epinephrine (r = 0.367; P = 0.0014) but did not correlate with age, BMI-SD or peak serum GH. Children with intact and abnormal GH responses presented similar mean peak serum cortisol levels (20.0 vs. 20.6 µg/dL/555 vs. 572 nmol/L; P = 0.21). CONCLUSION: Our data indicate that the current cut-off to define normal HPA axis response in children after insulin-induced hypoglycemia warrants reevaluation to avoid over-diagnosis of adrenal insufficiency. Our results suggest that peak serum cortisol levels ≥ 15.4 µg/dL (428 nmol/L) in children undergoing ITT might represent a normal cortisol response to stress, regardless of age, BMI or GH secretory capacity.

Investigation of the Effects of Trehalose on Glycemic Indices in Streptozotocin-Induced Diabetic Rats.

BACKGROUND AND AIM: Diabetes is a chronic metabolic disorder with considerable morbidity and mortality because of its associated complications that has become a challenging health problem worldwide. Trehalose (mycose) is a nonreducing disaccharide with a unique therapeutic potency without adverse effects, which has been found to improve glucose metabolism and homeostasis in different diabetes models. We hypothesized that trehalose can reduce blood glucose and improve insulin sensitivity. We have conducted this study to evaluate the effect of trehalose on glycemic indices in streptozotocin (STZ)-induced diabetic rats. METHOD: Fourteen diabetic rats were randomly assigned in two treatment groups (seven rats per group) that received trehalose at a dose of 1.5 g/kg/day via oral gavage and a dose of 45 mg/kg/day via intraperitoneal (i.p.) injection. Three control groups, including a positive control, diabetic control (DC), and nondiabetic rats as a normal control group (NC), received metformin (200 mg/kg/day), normal saline, and citrate buffer, respectively. The levels of fasting blood glucose (FBG) were measured at baseline (week 0) and after 4 weeks of treatment. Moreover, an oral glucose tolerance test (OGTT) was performed at the end of the study to determine glucose tolerance. RESULTS: The results showed that FBG levels were significantly decreased by -66% (-221 ± 65 mg/dL, p = 0.01), -40% (-114 ± 46 mg/dL, p = 0.02), and - 72% (-191 ± 68 mg/dL, p = 0.01) in trehalose-oral, trehalose-i.p., and metformin groups, respectively, after 4 weeks of administration. Evaluating the results of glucose tolerance test and analysis of corresponding areas under the glucose curve (AUCglucose) over 180 min indicated that glucose tolerance was significantly improved in the trehalose-i.p. group (p = 0.03) compared to DC group. CONCLUSION: Our findings suggested that trehalose administered via i.p. route might reduce FBG levels and improve glycemic control in STZ-induced diabetic rats.

Cortisol cut-points for the glucagon stimulation test in the evaluation of hypothalamic pituitary adrenal axis.

Diagnosis of adrenal insufficiency requires evaluation by dynamic stimulation tests. The insulin tolerance test (ITT) is accepted as the gold-standard test for the evaluation of hypothalamo-pituitary-adrenal (HPA) axis but the test is unpleasant and dangerous. Although it takes more time, glucagon stimulation test (GST) is a good alternative to ITT. The primary aim of this study was to compare the ITT and GSTs in the evaluation of HPA axe in patients with pituitary disorders. We conducted a prospective study in which ITT and GST were performed within 7 days in 81 patients. Serum cortisol was measured. We divided our population in Group 1 (G1): Adrenal Insufficiency (Peak cortisol under ITT <200 ng/mL) and Group 2 (G2): normal response (Peak cortisol under ITT >200 ng/mL). Receiver-operating characteristic (ROC) analysis was performed to identify the thresholds for GST. The mean peak of cortisol under GST was not significantly different from that obtained after ITT in the whole cohort (182.67 ± 89.07 ng/mL vs. 179.75 ± 79.01 ng/mL), and it was significantly reduced in patients of G1 (p < 10-3). ROC curve analysis showed that the best diagnostic accuracy was obtained with a peak cortisol cut-off to GST of 167 ng/mL (sensitivity, 89%; specificity, 79%). Using this cut-off, 86.4% of the patients were correctly classified. In our prospective series, GST is a potential accurate and safe alternative test for the assessment HPA. Test-specific cut-offs should be applied to avoid misinterpretation.

Hypopituitarism 3 and 12 months after traumatic brain injury and subarachnoid haemorrhage.

OBJECTIVE: High prevalence of hypopituitarism (HP) has been reported after traumatic brain injury (TBI) and subarachnoid haemorrhage (SAH). The aim of the study was to prospectively evaluate the prevalence and progression of HP in patients after TBI and SAH in Icelandic population. DESIGN: A 12 month prospective single-centre study. METHODS AND PROCEDURES: A total of 27 patients were included, 15 patients with TBI and 12 patients with SAH. Pituitary function was evaluated with baseline hormone measurements and diagnostic tests. An insulin tolerance test was used unless contraindicated, then the GHRH-arginine test and Synachten test were used. RESULTS: At three months, 16.7% (2/12) of the patients had HP after TBI and 33.3% (4/12) after SAH. At 12 months, 21.4% (3/14) of patients had HP after TBI and 9.1% (1/11) after SAH. Gonadotropin deficiency was the most common deficiency at 3 months and GH and gonadotropin deficiency at 12 months. CONCLUSIONS: There is a considerable risk of HP after TBI and reason to study pituitary function further in patients with SAH. We believe that neuroendocrine evaluation is important in these patients. Since recovery commonly occurs 12 months after the event, evaluation should be performed after that time if not clinically indicated earlier.

Effect of maternal dyslipidaemia on the cardiorespiratory physiology and biochemical parameters in male rat offspring.

The present study evaluated the effects of maternal dyslipidaemia on blood pressure (BP), cardiorespiratory physiology and biochemical parameters in male offspring. Wistar rat dams were fed either a control (CTL) or a dyslipidaemic (DLP) diet during pregnancy and lactation. After weaning, both CTL and DLP offspring received standard diet. On the 30th and 90th day of life, blood samples were collected for metabolic analyses. Direct measurements of BP, respiratory frequency (RF), tidal volume (VT) and ventilation (VE) under baseline condition, as well as during hypercapnia (7 % CO2) and hypoxia (KCN, 0·04 %), were recorded from awake 90-d-old male offspring. DLP dams exhibited raised serum levels of total cholesterol (TC) (4·0-fold), TAG (2·0-fold), VLDL+LDL (7·7-fold) and reduced HDL-cholesterol (2·4-fold), insulin resistance and hepatic steatosis at the end of lactation. At 30 d of age, the DLP offspring showed an increase in the serum levels of TC (P<0·05) and VLDL+LDL (P<0·05) in comparison with CTL offspring. At 90 d of age, DLP offspring exhibited higher mean arterial pressure (MAP, approximately 34 %). In the spectral analysis, the DLP group showed augmented low-frequency (LF) power and LF:high-frequency (HF) ratio when compared with CTL offspring. In addition, the DLP animals showed a larger delta variation in arterial pressure after administration of the ganglionic blocker (P=0·0003). We also found that cardiorespiratory response to hypercapnia and hypoxia was augmented in DLP offspring. In conclusion, the present data show that maternal dyslipidaemia alters cardiorespiratory physiology and may be a predisposing factor for hypertension at adulthood.

Factors associated with an inadequate hypoglycemia in the insulin tolerance test in Japanese patients with suspected or proven hypopituitarism.

We attempted to identify the predictors of an inadequate hypoglycemia in insulin tolerance test (ITT), defined as a blood glucose level higher than 2.8 mmol/L after insulin injection, in Japanese patients with suspected or proven hypopituitarism. A total of 78 patients who had undergone ITT were divided into adequate and inadequate hypoglycemia groups. The relationships between the subjects' clinical parameters and inadequate hypoglycemia in ITT were analyzed. Stepwise logistic regression analysis identified high systolic blood pressure (SBP) and high homeostasis model assessment of insulin resistance (HOMA-IR) as being independent factors associated with inadequate hypoglycemia in ITT. Receiver operating characteristic (ROC) curve analysis revealed the cutoff value for inadequate hypoglycemia was 109 mmHg for SBP and 1.4 for HOMA-IR. The areas under ROC curve for SBP and HOMA-IR were 0.72 and 0.86, respectively. We confirmed that high values of SBP and HOMA-IR were associated with inadequate hypoglycemia in ITT, regardless of the degree of reduction of pituitary hormone levels. Furthermore, the strongest predictor of inadequate hypoglycemia was obtained by using the cutoff value of HOMA-IR. Our results suggest that HOMA-IR is a useful pre-screening tool for ITT in these populations.

Algorithm for Screening of Adrenal Function in Stable Patients with Cirrhosis.

INTRODUCTION AND AIMS: Adrenal insufficiency (AI) is common in patients with cirrhosis. We aimed to assess the presence of AI in stable patients with cirrhosis using the gold-standard insulin tolerance test (ITT) and to propose an algorithm for screening AI in these patients. MATERIAL AND METHODS: We studied 40 stable patients with cirrhosis. We determined the basal total (BTC) and peak cortisol (PTC) levels. Using the ITT, we defined AI as a serum PTC < 18 µg/dL at 30 min after insulin-induced hypoglycemia. We assessed the diagnostic accuracy of BTC in different stages of liver disease to discriminate between those with NAF and AI. RESULTS: Of the 40 patients, 24 (60%) presented with AI. Child-Pugh and MELD scores differed between the NAF and AI groups (Child-Pugh: NAF 7.2 ± 1.7 vs. AI 8.8 ± 2.4, p = 0.024 and MELD: NAF 9.9 ± 2.5 vs. AI 14.9 ± 6.3, p = 0.001). The BTC level was lower in patients with AI than in those with NAF (7.2 ± 2.4 vs. 12.5 ± 5.2, p < 0.001). A BTC value ≤ 10.0 µg/dL had a 96% sensitivity (negative predictive value: 90%) for identifying AI. This cutoff value (BTC ≤ 10.0 µg/dL) provided 100% specificity (positive predictive value: 100%) in patients with advanced liver disease (Child-Pugh ≥ 9 or MELD ≥ 12). CONCLUSION: An algorithm including the use of BTC and the severity of liver disease may be a useful and simple method for assessing adrenal function in stable patients with cirrhosis.

Exogenous citrate impairs glucose tolerance and promotes visceral adipose tissue inflammation in mice.

Overweight and obesity have become epidemic worldwide and are linked to sedentary lifestyle and the consumption of processed foods and drinks. Citrate is a metabolite that plays central roles in carbohydrate and lipid metabolism. In addition, citrate is the additive most commonly used by the food industry, and therefore is highly consumed. Extracellular citrate can freely enter the cells via the constitutively expressed plasma membrane citrate transporter. Within the cytosol, citrate is readily metabolised by ATP-citrate lyase into acetyl-CoA - the metabolic precursor of endogenously produced lipids and cholesterol. We therefore hypothesised that the citrate ingested from processed foods and drinks could contribute to increased postprandial fat production and weight gain. To test our hypothesis, we administered citrate to mice through their drinking water with or without sucrose and monitored their weight gain and other metabolic parameters. Our results showed that mice receiving citrate or citrate+sucrose did not show increased weight gain or an increase in the weight of the liver, skeletal muscles or adipose tissues (AT). Moreover, the plasma lipid profiles (TAG, total cholesterol, LDL and HDL) were similar across all groups. However, the group receiving citrate+sucrose showed augmented fasting glycaemia, glucose intolerance and the expression of pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6 and IL-10) in their AT. Therefore, our results suggest that citrate consumption contributes to increased AT inflammation and altered glucose metabolism, which is indicative of initial insulin resistance. Thus, citrate consumption could be a previously unknown causative agent for the complications associated with obesity.

A Comparison of Salivary Steroid Levels during Diagnostic Tests for Adrenal Insufficiency.

Numerous diagnostic tests are used to evaluate the hypothalamic-pituitary-adrenal axis (HPA axis). The gold standard is still considered the insulin tolerance test (ITT), but this test has many limitations. Current guidelines therefore recommend the Synacthen test first when an HPA axis insufficiency is suspected. However, the dose of Synacthen that is diagnostically most accurate and sensitive is still a matter of debate. We investigated 15 healthy men with mean/median age 27.4/26 (SD±4.8) years, and mean/median BMI (body mass index) 25.38/24.82 (SD±3.2) kg/m2. All subjects underwent 4 dynamic tests of the HPA axis, specifically 1 µg, 10 µg, and 250 µg Synacthen (ACTH) tests and an ITT. Salivary cortisol, cortisone, pregnenolone, and DHEA (dehydroepiandrosterone) were analysed using liquid chromatography-tandem mass spectrometry. During the ITT maximum salivary cortisol levels over 12.5 nmol/l were found at 60 minutes. Maximum cortisol levels in all of the Synacthen tests were higher than this; however, demonstrating that sufficient stimulation of the adrenal glands was achieved. Cortisone reacted similarly as cortisol, i.e. we did not find any change in the ratio of cortisol to cortisone. Pregnenolone and DHEA were higher during the ITT, and their peaks preceded the cortisol peak. There was no increase of pregnenolone or DHEA in any of the Synacthen tests. We demonstrate that the 10 µg Synacthen dose is sufficient stimulus for testing the HPA axis and is also a safe and cost-effective alternative. This dose also largely eliminates both false negative and false positive results.

The Response of C19 Δ5-steroids to ACTH Stimulation and Hypoglycemia in Insulin Tolerance Test for Adrenal Insufficiency.

Studies on the time course of ACTH- or insulin-induced hypoglycemia stimulating adrenal androgens are usually limited to dehydroepiandrosterone and/or its sulphate. Our data on dehydroepiandrosterone (DHEA) and its hydroxylated metabolites clearly show that measurements of DHEA and its sulphate (DHEAS) are valuable markers of the integrity of the HPA (hypothalamus-pituitary-adrenal) axis. Assessments of HPA function should rely on measurements of baseline and/or stimulated serum cortisol concentrations, and C19 Δ5-steroids may provide additional information. The art of stimulation of 7- and 16-hydroxylated metabolites of DHEA can help our understanding of the formation sequence of these compounds.